Steroid hormone action is mediated by highly specific receptor proteins found in hormonally responsive tissues. However, to date there has been relatively little work on the large scale isolation and detailed molecular characterization of steroid hormone receptors from mammalian tissue. The objective of this study is to isolate and characterize the glucocorticoid receptor from a hormonally sensitive target organ, the lactating mammary gland. Our preliminary studies have shown that lactating goat mammary tissue is a rich source of this protein. We will develop purification procedures taking advantage of previous work by other investigators on the purification of steroid hormone receptors from various tissue sources. The purified glucocorticoid receptor will be characterized physico-chemically and the topography of the steroid binding site will be investigated by the use of radio-active affinity-labeling steroid derivatives and site specific protein modifying agents. The purified receptor will also be used for the production of an antiserum that may be helpful in the development of sensitive immunoassays to detect the glucocorticoid receptor in biopsy specimens from patients with certain forms of hormonally responsive cancer. The "activated" form of the receptor will be purified and the mechanism of receptor "activation" will be studied. Binding of "activated" receptor to DNA-cellulose, isolated nuclei and nuclear subfractions will be investigated and the effect of modification of specific amino acid residues in the receptor on nuclear binding will be determined. These studies should help not only to clarify the nature of nuclear binding but also to identify amino acid residues in the nuclear attachment site of the receptor protein.